Class I MHC (MHC-I) molecules primarily present endogenous antigens, i.e. antigens that are present in the cytosol and are subject to the cytosolic processing mechanisms that comprise the conventional MHC-I processing pathway. However, exogenous antigens can also be presented by MHC-I molecules under certain circumstances, particularly in the case of particulate antigens. Recently, considerable attention has been focused on mechanisms that may contribute to alternate MHC-I processing pathways. Divergent results in several different systems have suggested that more than one alternate processing mechanism may exist. In some cases, MHC-I molecules present vacuolar antigens via alternate MHC-I processing mechanisms that are quite distinct from the conventional MHC- I processing pathway. These mechanisms may play important roles in generating CD8 T cell responses, especially to antigens expressed by vacuolar microorganisms and tumor cells. This project will define the mechanisms involved in the alternate MHC-I processing of several different types of exogenous antigens (intravacuolar bacteria and particulate vaccines). These studies may elucidate the basis of CD8 T cell responses to intravacuolar pathogens, such as Leishmania, Salmonella, and Toxoplasma species, and Mycobacterium tuberculosis. These studies also apply to tumor immunology and the role of alternate MHC-I processing mechanisms in the genesis of anti-tumor CD8 T cell responses. In addition, these studies will provide basic information that may be applied to develop strategies of therapeutic immunization to achieve protective CD8 T cell responses with non-viable vaccine preparations, in the absence of the endogenous antigen synthesis that is provided by live viral vaccine preparations. The use of non-viable particulate vaccines would be a safer way to elicit CD8 immunity in immunocompromised patients.